Platelet-derived growth factor (PDGF) -BB is a potent chemoattractant for mesenchymal cells. Intracellular signal transduction for PDGF-indeuced chemotactic response has been reported to be dependent on phosphatidylinositol 3-kinase (P13K) actication. Here we report a P13K-independent pathway operating for PDGF-induced chemotaxis in vascular smooth musclw cells (SMCs) and other cell types. Two different P13K inhibitors, wortmannin (WT,1nM-1 FM) and LY294002 (100nM-10 FM) did not inhibit PDGF-induced chemotaxis in SMCs and Swiss 3T3 cells, whereas WT inhibited activity of P13K that were immunopurified from PDGF-stimulated cells as well as P13K purified from cells which were stimulated with PDGF in the presence of the same concentrations of WT.Similarly, WT (100 nM) abolished the increase in intracellular phosphatidylinositol (3,4,5) triphosphate after PDGF stimulation. Furthermore, CHO/)p85 cells overexpressing a dominant negative p85 subunit of P13K showed a chemotactic response comparable to that of parental cells, while showing a remarkable decrease in P1K activity. Rapamycin, a specific inhibitor of pp70 S6 kinase, which is one of the well characterized-downstreams of P13K,didnot inhibit PDGF-induced chemotaxis. Both WT and LY294002 inhibited PDGF-induced amino acid uptake and actin-stress fiber reorganization, and partly inhibited PDGF-induced lucose incorporation in Swiss 3T3 cells. Our findings indicate that, in vascular smooth muscle cells and other cell types, the signal transduction for PDGF-induced chemotaxis is independent of P13K activity, while the signal transduction for PDGF-induced amino acid uptake, glucose incorporation and cytoskeletal reorganization are dependent on P13K.